PROJECT SUMMARY/ABSTRACT The goal of this proposal is to determine how the ?canonical to non-canonical? WNT signaling shift contributes to emphysema and impaired lung regeneration. Emphysema is a chronic lung disease affecting over 4 million people in the US and therapies that stop the progression or reverse disease are not available. Emphysema is characterized by progressive and permanent destruction of parenchymal lung tissue, in particular functional alveolar epithelium. Importantly, the endogenous ability of the distal lung to activate self-repair mechanisms is defective in emphysema. Distal lung epithelial progenitor cell (DLEP) populations that might contribute to lung repair have been identified in the human lung, which raises the question why and which regenerative pathways and/or cells are silenced in emphysema. Canonical WNT/?-catenin signaling has been recently identified as a potential regenerative pathway with reduced activity in emphysema. Notably, non-canonical WNT signaling, which is ?-catenin independent, can inhibit canonical WNT/?-catenin signaling. We recently reported increased non-canonical WNT signaling in emphysema, representing the first time a canonical to non-canonical WNT signal shift has been demonstrated to contribute to a chronic lung disease. In human and experimental emphysema, the non-canonical WNT ligand WNT5A is increased. Inhibition of WNT5A restored canonical WNT/?-catenin signaling in alveolar epithelial cells in vitro and attenuated tissue destruction in two emphysema models in vivo. Here, we aim to identify the molecular mechanisms and functional consequence of the inhibitory effect of WNT5A on canonical WNT/?-catenin signaling in DLEP and lung regeneration in emphysema. We hypothesize that non- canonical WNT signaling, mediated by disease- and cell-specific expression of WNT receptors, acts as a ?brake? for endogenous WNT/?-catenin-driven lung regeneration in emphysema. Aim 1: determine whether and how WNT/?-catenin-responsive (WNT)-DLEP might change in emphysema and how WNT5A impacts WNT-DLEPs, using lung organoid assays complemented by in vivo and ex vivo studies in healthy and emphysematous lung tissues. Aim 2: identify the non-canonical WNT receptor(s) on DLEP that mediates WNT5A signaling and test the hypothesis that these are potential therapeutic targets to restore WNT signal balance and thereby attenuate experimental emphysema in vitro and in vivo. Aim 3: determine whether reversing the canonical to non-canonical WNT signal shift accelerates lung regeneration in experimental emphysema in vivo and leads to a decrease in COPD/emphysema biomarkers in patient-derived 3D-LTCs ex vivo. This proposal will thus 1) provide in-depth knowledge about novel WNT/?-catenin-responsive DLEP subpopulations and be the first to elucidate the role of non-canonical WNT signaling for DLEP function in emphysema; 2) identify cell-specific WNT receptors in the healthy and diseased lung, an area largely unexplored in lung pathologies and critical for development of novel therapeutic targets; and 3) result in significant proof-of-principle data from patient-derived tissue to further pave the way for future translational and clinical investigations in emphysema.